downloadfile.PDF

Introduction to Medical Microbiology (MB 241) DR:Abir Aburas  READING MATERIALS Handout by lecturer Jawetz’s Medical Microbiology Murray  READING MATERIALS Immunology lecture notes ( AL-TUBULY AA and DAEKI AO) Immunology le...

Introduction to Medical Microbiology (MB 241) DR:Abir Aburas  READING MATERIALS Handout by lecturer Jawetz’s Medical Microbiology Murray  READING MATERIALS Immunology lecture notes ( AL-TUBULY AA and DAEKI AO) Immunology lecture notes (Prof. Salam Al-Geali )  Course Objectives This course covers principles of microbiology microbial structure and function. Pathogenesis. Classification and host-pathogen relationship. Control of pathogens. Disease transmission, host resistance, and immunity.  Definitions Microbiology: The study of microorganisms(viruses, bacteria, algae, fungi, and protozoa) too small to be seen with the naked eye. Microbe (microorganism): are minute living things that are usually unable to be viewed with the naked eye. some are pathogenic and many are beneficial. Microbiologists: Are scientists who studies the characteristics and functions such as morphology, cytology, physiology, ecology, taxonomy, genetics, and molecular biology.  Medical microbiology Definition : is the study of causative agents of infectious diseases to humans. Bacteriology : is the study of bacteria, the causative agents of a member of infectious diseases Virology : is the study of viruses, non-cellular living systems, capable of causing infectious diseases in man Mycology : is the study of fungi pathogenic for man  Protozoology : is the science that deals with pathogenic unicellular animal organisms Parasitology : is study of parasites (protozoa and helminths) Immunology : is the study which concerned with mechanisms of body protection against pathogenic microorganisms and foreign cells and substances  Importance of Microbiology Microbes make the Universe, there are > 5 x 1030 types Microbes in the world. Humans have intimate relation with Microbes > 90% of the cells in our Body are Microbes. Only a minority are pathogenic. Most of our problems are caused by microbes. Some industrial uses of microbes ( food processing, antibiotics, Human Insulin, human growth factor…etc. Microbes in research (Easy to grow, Simple and easy to.study)  Emerging infectious diseases EID’s New evolutionary features (virulence and resistance) Response to man violating the environment (Ebola virus, SARS) Weapons of mass destruction  History of Microbiology van Leeuwenhoek (1632–1723): was the first microbiologist and the first person to observe bacteria using a single-lens microscope of his own design.( father of microbiology) Louis Pasteur (1822–1895): Pasteur developed a process (today known as pasteurization) to kill microbes. pasteurization is accomplished by heating liquids to 63° to 65°C for 30 minutes or to 73° to 75°C for 15 seconds. Robert Koch (1843–1910): was a pioneer in medical microbiology and worked in cholera, anthrax and tuberculosis. He was awarded a Nobel prize in 1905 (Koch's postulates) he set out criteria to test. Alexander Fleming (1929): Discovered penicillin  History of Microbiology 1590 – First compound light microscope (Zacharias Janssen )  1676 –first observation of bacteria “animalcules” Anton Von Leeuwenhoek  1796 – First vaccine (smallpox) Edward Jenner  1867 Joseph Lister Practiced Antiseptic Surgery Discovered carbolic acid to kill microorganisms and surgical dressings  1884 Koch’s Postulates of Disease Transmission Robert Koch Father of bacteriology Formulated the bacteriological techniques Staining methods Discovered the mycobacterium and vibrio cholera  1857 – Germ Theory of Disease Louis Pasteur Contributed best in microbiology sterilization Hot Air oven Autoclave  1885 -Louis Pasteur Coined the word Vaccine Vaccine against Rabies Anthrax vaccine Considered the father of Modern Microbiology  1929 Discovery of Penicillin (first antibiotic) (Alexander Fleming)  John Tyndall 1820-1893 Discovered highly resistant bacterial structure ( Endospore) in air and soil. Tyndallisation : (fractional sterilization) prolonged boiling or intermittent heating was necessary to kill endospore  1938 – First Electron Microscope (is capable of magnifying biological specimens up to one million times.  1953 Structure of DNA Revealed Watson & Crick  1954 Polio Vaccine Jonas Salk  Recent History Genetic engineering Cloning Human Genome Project Biotechnology Basis for classification and laboratory diagnosis of microorganisms DR.Abir ABURAS  Bacterial classification  19th century, all organisms were divided into two kingdoms plant and animal  In 1866, a third kingdom, the Protista  In 1925, four kingdom system, differentiate between prokaryotes and eukaryotes within the Protista kingdom  In 1969, H.R. Whittaker, five kingdom classification  Diversity of Microbes  Bacteria- single celled prokaryotes, Peptidoglycan cell walls, Binary fission.  Protozoa- eukaryotic, single celled, colonial, many ways of nutrition, motile via pseudopods, cilia, or flagella.  Fungi- absorb nutrients, single celled filamentous.  Viruses- A cellular, DNA or RNA core , surrounded by a protein coat , enclosed in a lipid envelope , replicated only when they are in a living host cell.  Algae: Eukaryotes, cellulose cell walls, use photosynthesis for energy.  Archaea: Prokaryotic, Lack peptidoglycan, Live in extreme environments (Methanogens, Extreme halophiles,Extreme thermophiles  Others- worms, insects  Classification Systems Taxonomy system, classification of living organisms into groups: 1) Phylogenetic Classification System: groups reflect genetic similarity and evolutionary relatedness 2) Phenotyping Classification System: groups based on observable characteristics  Levels of Classification Taxonomy  A group or “level” of classification  Hierarchical; broad divisions are divided up into smaller divisions: Kingdom: Eubacteria Phylum: Proteobacteria Class: Gamma proteobacteria Order: Enterobacteriales Family: Enterobacteriaceae Genus: Escherichia Species: Escherichia coli Strain: E. coli (biovar or morphovar) Type: E. coli O157 Nomenclature  Scientific name (Systematic Name) Binomial System of Nomenclature  Genus name + species name 1. Italicized or underlined 2. Genus name is capitalized and may be abbreviated 3. Species name is never abbreviated 4. A genus name may be used alone to indicate a genus group; a species name is never used alone 5. eg: Bacillus subtilis B. subtilis  Kingdom of bacteria 1) Archaebacteria (Ancient bacteria)  Found in harsh environments  It Lack peptidoglycan in cell walls  Different lipids in their cell membrane  Different types of ribosomes  Different gene sequences  subdivided into 3 groups: a. Methanogens b. Halophiles c. Thermoacidophiles or Thermophiles  Kingdom of bacteria 2) Eubacteria (true bacteria)  Most bacteria are in this group  Have three basic shape (coccus, bacillus, spiril)  May be aerobic or anaerobic  Identified by Gram staining  Kingdom of bacteria 3) Cyanobacteria  Gram negative  photosynthetic  Called blue-green bacteria  Contain phycocyanin (red-blue) pigments & chlorophyll  Criteria for classification of bacteria 1) Microscopically:  Cell shape or morphology (cocci, bacilli, spiral, coma & clubed)  Arrangement of bacteria (clusters, chains, diplococci)  Specialized structure (spore or flagella)  Spherical (cocci) bacteria  Micrococci  Diplococci  Streptococci  Staphylococci  Tetracocci  Sarcine  Criteria for classification of bacteria 2) Staining procedures  Gram negative bacteria  Gram positive bacteria  Acid fast bacteria  Lack staining (lack cell wall)  Criteria for classification of bacteria 3)Colony morphology  Different bacteria have different colonial characteristics  Zone of hemolysis in agar medium containing blood agar  Some bacteria produce pigments and enzymes that can be detected zones surrounding colonies  Criteria for classification of bacteria 4)Biochemical characteristics bacterial enzymes to identify the bacteria and distinguish between bacterial species. Urease test: some bacteria produce urease enzyme  Criteria for classification of bacteria 5)Growth characteristics  Oxygen requirement 1)Aerobic 2)Anaerobic 3)Microaerophilic, aerotolerant 4)Facultative anaerobic  Fastidious/non-fastidious  Criteria for classification of bacteria 6) Antigenic characteristics  Serological test (using antisera to detect unknown organism)  Based on Ag-Ab reaction 7) Genetic characteristics  G + C content  Nucleic acid sequencing  Major categories and groups of bacteria  four major categories of bacteria based on character of cell wall into: 1) Gram-negative eubacteria : have cell wall and grouped into (16 groups) based on differences on several characters such as shape, motility, reproduction, respiratory & living intra or extra cellular. Ex: Treponema sp, Campylobacter sp, Brucella sp, 2) Gram-positive eubacteria : have cell wall and grouped into 6 groups based on shape, spore, respiration. Examples: Staphylococci, Clostridium, Listeria, Actinomyces, 3) Eubacteria lacking cell wall (one group) such as Mycoplasma. 4) Archaebacteria (5 groups)  Laboratory diagnostic microbiology Microscopic examination of specimen ( stained or non stained) Culture and biochemical tests Serological tests Molecular diagnosis Immunological assays  Microscopic examination Staining of Bacteria used for study the shape and arrangement of bacteria. There are two types of dyes: Basic dye: positively charged and can combined with bacterial negative charge.  Acidic dye: do not stain bacterial cell and use as background material contrasting color.  Types of Stains I. Simple Stain iii. Special stain II. Differential stain  Capsule stain  Gram Stain  Spore stain  Acid Fast Stain  Negative satin  Flagella stain I. Simple stain  Stain same color to all bacteria.  Retain bacterial morphology and arrangement.  e.g. Methylene blue, Carbol fuchsin , Safranine, etc II. Differential stain 1- Gram stain:  Developed in 1884 by Hans Gram  It differentiates bacteria into two broad groups :  Gram positive appear violet  Gram negative appear red  Used purple Crystal Violet & red Safranin stains  G-ve bacteria loss crystal violet stain due to LPS and very thin peptidoglycan layers 2- Acid Fast Stain (Ziehl & Neelsen STAIN)  For staining of mycobacteria & some Actinomyces.  They have high content of Lipids (mycolic acid) Fatty acid in their cell wall.  Carbol fuchsin is primary stain & decolorization by acid alcohol. III. Special stain 1- Negative stain  Produce colored background against which bacteria stand out in contrast.  Used for bacterial capsule, spirochetes. e.g. India ink, Nigrosin 2- spore staining 3- capsular staining 4- flagella staining  Culture media  Food that is used for culturing microorganisms is called Media.  Bacterial culture consists of solution or suspension of nutrients that may be solidified by agar.  Agar is chemical from seaweed that melts at 100C and freezes at 45C  The main nutrients including: Carbon source Nitrogen source Electrolytes Minerals  Culture Media state  Two basic culture techniques used in microbiology:  Broth - a liquid medium – prepared in culture tubes (test tubes) – used for pure culture of bacteria, algae and some fungi  Agar : 1. a semi solid medium (for motility) 2. solid medium (for isolation – colonial study)  Types of culture media  Simple media (Basal media)  Specialized media  Synthetic media  Aerobic and anaerobic media  Cell culture for obligate intracellular bacteria (e.g., Chlamydia spp) I-Simple Media: It contains an essential requirement. Ex. Nutrient media, Nutrient broth, II- Specialized Medium 1) Transport Medium Not a true medium; used to transport samples to the lab. 2) Enriched Medium more” nutritious ingredient is added such as blood, serum, egg yolk, proteins, vitamins, …etc. Example: blood agar, chocolate agar III. Selective Media  Allows the growth of some organisms but inhibits the growth of others  Examples: Eosin Methylene Blue (EMB) inhibits Gram-positives therefore selects for Gram-negatives. It is selective and differential medium used for Isolation of fecal coliform.( normal gut flora) Lowenstein Jensen media (Mycobacterium tuberculosis) IV. Differential Media  allows the differentiation between 2 groups of bacteria  EXAMPLE: Blood Agar Medium- can distinguish between 1) Alpha-Hemolysis- incomplete RBC lysis 2) Beta Hemolysis- complete RBC lysis 3) Gamma Hemolysis- no RBC lysis ( Streptococcus pyogenes) Selective & differential Media Mannitol salt agar for Staphylococcus aureus v. Enrichment Broths(media)  encourage the growth of a particular type of microbe.  Example: Selenite F broth- enriches for Salmonella & Shigella.  Anaerobic Culture Methods  Reducing media  EXAMPLES: 1) Anaerobic jar 2) Anaerobic chamber  Identification of bacteria Catalase test  Oxidase test  Coagulase test  Urease test EPI kit  Serological methods  Detection of microbial antigen with known antiserum.  Quellung reaction: Some bacteria having capsules can be identified directly in clinical specimens by a reaction that occurs when the organisms are treated with serum containing specific antibodies (S. pneumonia).  Molecular Diagnostics  DNA Sequencing  PCR  Restriction endonuclease analysis  RAPD (random amplified polymorphic DNA)  DNA fingerprinting